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Some of Our Group's Previous Work - Squeezing Viruses with an Atomic Force Microscope

We know from our studies with lambda phage that viral capsids can support internal pressures of 50 - 60 atm. The interactions between the proteins that make up the capsid are held together by hydrophobic and electrostatic forces and hydrogen bonds. How can structures that are joined by relatively weak bonds be so strong? Just how strong are viral capsids? Are phage capsids, which are [l|phage_packaging|highly pressurized], stronger than capsids of plant viruses like CCMV, do not have to support high pressures? We are answering such questions by nanoindentation experiments in which we measure the forces required to deform viral capids. The work is being carried out with our collaborators at the Free University of Amsterdam.

The capsids are adsorbed on a hydrophobic glass surface and then studied with an Atomic Force Microscope (AFM) in a buffer solution. After individual capsids are imaged - see left - the tip of the AFM, which is attached to a flexible cantilever, is positioned above the center of the capsid and the capsid is then compressed between it and the supporting glass surface. The force on the tip and its displacement are measured throughout the compression. Typical plots of force (in nanoNewtons, nN) vs displacement (in nanometers, nm) are shown below, right, for CCMV. To analyze the curves we have to recognize that the cantilever and the capsid are like two springs in series and therefore subjected to the same force. The black line in the figure represents the bending of the cantilever when it is pushed against an incompressible surface (the glass next to the capsid); the red lines are repeated compressions of the capsid. At each force, the distance between the black line and the force curve for the capsid represents the decrease in the capsid diameter.

In our early measurements we have found that, remarkably, empty capsids behave completely elastically for compressions of up to 20% at forces up to 1 nN. Although they have a complex protein structure, to a first approximation, the capsids act like uniformly thin shells of an elastic material and their response to compression is similar to that of hard plastics. The mechanical behavior of more realistic models of capsid structure can be analyzed by the method of finite element analysis; our collaborator Professor [story:klug Bill Klug] of the UCLA Mechanical and Aerospace Engineering Department is examining this problem. When the compression force exceeds about 1 nN, the capsids undergo irreversible breakage, as seen in the figure above, left.

We are also studying how the mechanical properties of the shells change when they contain their RNA genome. There are many mutants of CCMV with known structures, allowing us to examine the effects of changes in primary protein sequences and the interactions between amino acid residues on the capsid strength. The interactions between the proteins in the shell change with pH and the CCMV capsid undergoes a reversible radial expansion at pH 7.5. Preliminary results show that the pH change produces a major reduction in the capsid stiffness and extends the range of elastic behavior to compressions of 50%.